Cu appears to have many important functional roles in the body that apparently relate, among others, to the maintenance of immune function, bone health and haemostasis. Some have suggested a role for long-term marginal Cu deficiency in the aetiology of a number of degenerative diseases. Accurate diagnosis of marginal Cu deficiency, however, has remained elusive despite an increased understanding of the biochemistry of Cu and its physiological roles in the body. Traditional markers of Cu status, such as serum Cu and caeruloplasmin protein concentrations are insensitive to subtle changes in Cu status. Cu-containing enzymes, such as Cu–Zn-superoxide dismutase, cytochrome c oxidase and diamine oxidase, may be more reliable but evidence to date is not conclusive. Development of markers sensitive to marginal Cu status is essential before conclusions can be drawn concerning the risks of long-term intake of suboptimal dietary Cu. As Cu appears to be essential for maintenance of immune function, activities of specific immunological markers, altered in Cu deficiency, offer alternatives. This review evaluates a selection of immunological markers that could be considered potentially sensitive markers of marginal Cu status. The indices of immune function reviewed are neutrophil function, interleukin 2 production, blastogenic response to mitogens and lymphocyte subset phenotyping.

Diet, among other environmental and genetic factors, is currently recognised to have an important role in health and disease. There is increasing evidence that the human colonic microbiota can contribute positively towards host nutrition and health. As such, dietary modulation has been proposed as important for improved gut health, especially during the highly sensitive stage of infancy. Differences in gut microflora composition and incidence of infection occur between breast- and formula-fed infants. Human milk components that cannot be duplicated in infant formulae could possibly account for these differences. However, various functional food ingredients such as oligosaccharides, prebiotics, proteins and probiotics could effect a beneficial modification in the composition and activities of gut microflora of infants. The aim of the present review is to describe existing knowledge on the composition and metabolic activities of the gastrointestinal microflora of human infants and discuss various possibilities and opportunities for its nutritional modulation.

Tannins have received considerable attention from animal nutritionists as potential agents for modifying ruminal fermentation patterns, or for exploring new feed resources. This group of secondary plant compounds is defined by their ability to form complexes with proteins. A widely accepted method for assaying the biological activity of extracted tannins is the precipitation of bovine serum albumin. The protein carries a radioactive label (125I) to allow direct quantification from the precipitate. Tannin–protein complexes dissolve in sodium dodecylsulfate solution. A dot-blot assay for protein determination, which is based on the reversible binding of a fluorochrome, benzoxanthene yellow, to the protein spots and is not disturbed by the presence of detergents, can replace the radioactive method by a fluorimetric measurement. A novel alternative to the last part of the dot-blot assay is to scan the stained protein spots in situ using a video camera and computer image analysis. Several filter sets were tested and, within a concentration range of 0·1–2·0 mg protein/ml, each of them yielded results identical to the original method while the time required was only 30 % of the working time consumed by the original procedure. The modified dot-blot assay should be applicable to the evaluation of tannin activity in all shrub and tree foliages considered as animal feed.

This work was designed to investigate the effect of different lipid sources on ob gene expression and serum leptin levels in rats with two different feeding protocols: (1) free access to food; or (2) energy-controlled feeding. Male Wistar rats were fed diets containing 40 % energy as fat (olive oil, sunflower oil or beef tallow), for 4 weeks. In Expt 1 rats had free access to food, and in Expt 2 rats were fed a controlled amount of food (16 g/d, equivalent to 300 kJ/d). Insulin and leptin were determined by ELISA and ob mRNA by Northern blot. When rats had free access to food, ob mRNA levels were higher in animals fed either olive oil or sunflower oil than in those fed beef tallow. In marked contrast with feeding ad libitum, no differences were found among dietary fat groups in rats fed energy-controlled diets. When both feeding protocols were compared, free access to food induced an increased expression of ob mRNA in perirenal and/or epididymal adipose tissues from rats fed either olive oil or sunflower oil, but not from rats fed beef tallow. Dietary lipid type did not induce modifications in serum leptin concentrations. A tendency to higher serum leptin levels was observed more in rats with free access to food than in rats fed energy-controlled feeding. No differences were found in insulin levels. Dietary fat type importantly affects ob mRNA expression in rat white adipose tissue under hyperphagic conditions. Further study is needed in order to elucidate the mechanism underlying this effect.

Fish-oil supplementation can reduce circulating triacylglycerol (TG) levels and cardiovascular risk. This study aimed to assess independent associations between changes in platelet eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) and fasting and postprandial (PP) lipoprotein concentrations and LDL oxidation status, following fish-oil intervention. Fifty-five mildly hypertriacylglycerolaemic (TG 1·5–4·0 mmol/l) men completed a double-blind placebo controlled cross over study, where individuals consumed 6 g fish oil (3 g EPA+DHA) or 6 g olive oil (placebo)/d for two 6-week intervention periods, with a 12-week wash-out period in between. Fish-oil intervention resulted in a significant increase in the platelet phospholipid EPA (+491 %, P<0·001) and DHA (+44 %, P<0·001) content and a significant decrease in the arachidonic acid (-10 %, P<0·001) and γ-linolenic acid (-24 %, P<0·001) levels. A 30 % increase in ex vivo LDL oxidation (P<0·001) was observed. In addition, fish oil resulted in a significant decrease in fasting and PP TG levels (P<0·001), PP non-esterified fatty acid (NEFA) levels, and in the percentage LDL as LDL-3 (P=0·040), and an increase in LDL-cholesterol (P=0·027). In multivariate analysis, changes in platelet phospholipid DHA emerged as being independently associated with the rise in LDL-cholesterol, accounting for 16 % of the variability in this outcome measure (P=0·030). In contrast, increases in platelet EPA were independently associated with the reductions in fasting (P=0·046) and PP TG (P=0·023), and PP NEFA (P=0·015), explaining 15–20 % and 25 % of the variability in response respectively. Increases in platelet EPA+DHA were independently and positively associated with the increase in LDL oxidation (P=0·011). EPA and DHA may have differential effects on plasma lipids in mildly hypertriacylglycerolaemic men.

The aim of this study was to evaluate the effect of dietary oxysterols on coronary atherosclerosis and vasospasm. Golden Syrian hamsters were fed three diets with different lipid contents for 3 months: (1) a normolipidaemic diet containing 25 g corn oil–fish oil (4:1, w/w)/kg (group Low L); (2) a hyperlipidaemic diet composed of the normolipidaemic diet supplemented with 150 g lard+30 g cholesterol/kg (group High L); (3) a third diet, similar to the hyperlipidaemic diet, in which 4 g cholesterol/kg was replaced by a mixture of oxysterols (group High L+OS). The oxysterol mixture contained (g/kg): 5,6α-epoxycholesterol 211, 5,6β-epoxycholesterol 179, 7α-hydroxycholesterol 67, 7β-hydroxycholesterol (7βOH) 185, 7-ketocholesterol (7 K) 235; and trace amounts of 7-hydroperoxycholesterols (approximately 30 g/kg). Atherosclerosis was evaluated by measuring myocardial Ca, oxysterols and acyl-CoA cholesterol acyl transferase (ACAT) activity; furthermore, coronary reactivity to sodium nitroprusside (5×10-6 m) was measured and the morphology of coronary arteries was visualized by transmission electron microscopy. Coronary spasm was determined by evaluating reactivity to serotonin (5×10-6 m). Feeding the high-lipid diet (group High L) increased the plasma level of 7βOH, 7 K and cholestanetriol. The presence of oxysterols in the diet (group High L+OS) further increased the concentrations of 7βOH and 7 K in the plasma. However, as evidenced by myocardial Ca, ACAT activity and coronary reactivity to sodium nitroprusside, severe atherosclerosis did not develop during the 3-month diet. 7 K was increased in myocardial lipids of groups High L and High L+OS. Electron microscopy did not show the development of atherosclerosis in group High L, whereas vascular wall thickening, endothelial damage and smooth muscle cell proliferation and migration occurred when oxysterols were present in the food. Serotonin (5×10-6 m) induced exacerbated coronary vasoconstriction in group High L that was completely reversed by dietary oxysterols. In conclusion, dietary oxysterols exhibit anti-spasmodic properties, but they cannot be used as agents against excess dietary lipid-induced coronary spasm because of their atherogenic properties.

Besides being an indispensable amino acid for protein synthesis, arginine (Arg) is also involved in a number of other physiological functions. Available data on the quantitative requirement for Arg in different teleosts appear to show much variability. So far, there are very limited data on the maintenance requirements of indispensable amino acids (IAA) in fish. In the present study, we compared N and Arg requirements for maintenance and growth of four finfish species: rainbow trout (Oncorhynchus mykiss), turbot (Psetta maxima), gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax). Groups of fish having an initial body weight close to 5–7 g were fed semi-purified diets containing graded levels of N (0 to 8 % DM) and Arg (0 to 3 % DM) over 4 to 6 weeks. For each species, N and Arg requirements for maintenance and for growth were calculated regressing daily N gain against daily N or Arg intakes. N requirement for maintenance was estimated to be 37·8, 127·3, 84·7 and 45·1 mg/kg metabolic body weight per d and 2·3, 2·2, 2·6 and 2·5 g for 1 g N accretion, in rainbow trout, turbot, gilthead seabream and European seabass respectively. The four species studied appear to have very low or no dietary Arg requirements for maintenance. Arg requirement for g N accretion was calculated to be 0·86 g in rainbow trout and between 1·04–1·11 g in the three marine species. Turbot required more N for maintenance than the other three species, possibly explaining its reputedly high overall dietary protein requirement. Data suggest a small but sufficient endogenous Arg synthesis to maintain whole body N balance and differences between freshwater and marine species as regards Arg requirement. It is worth verifying this tendency with other IAA.

The uptake of α-tocopherol and β-carotene and their antioxidative effect in plasma and buccal mucosal cells after oral application in twelve subjects is demonstrated in our study. The effect on the antioxidative status was evaluated using a modified thiobarbituric acid-reactive substance (TBARS) method. As expected, the supplement of 134·2 mg α-tocopherol/d and 25 mg β-carotene/d for 7 d resulted in a significant increase of α-tocopherol and β-carotene concentration in plasma (P<0·05). In buccal mucosal cells, the concentration of β-carotene increased after supplementation (P<0·05), whereas the concentration of α-tocopherol remained constant. A decrease in TBARS (P<0·05) was found in buccal mucosal cells but not in plasma. In conclusion, an uptake of the supplemented antioxidants was detected in plasma and in buccal mucosal cells. There was significant change in β-carotene concentration and oxidative stress as measured using a modified TBARS test in buccal mucosal cells, but not in the plasma.

Controversy exists about the optimal amount and source of dietary carbohydrate for managing insulin resistance. Therefore, we compared the effects on insulin sensitivity (SI), pancreatic responsivity (AIRglu) and glucose disposition index (DI=SI×AIRglu) of dietary advice aimed at reducing the amount or altering the source of dietary carbohydrate in subjects with impaired glucose tolerance (IGT). Subjects were randomized to high-carbohydrate–high-glycaemic index (GI) (high-GI, n 11), high-carbohydrate–low-GI (low-GI, n 13), or low-carbohydrate–high-monounsaturated fat (MUFA, n 11) dietary advice, with SI, AIRglu and DI measured using a frequently sampled, intravenous glucose tolerance test before and after 4 months treatment. Carbohydrate and fat intakes and diet GI, respectively, were: high-GI, 53 %, 28 %, 83; low-GI, 55 %, 25 %, 76; MUFA, 47 %, 35 %, 82. Weight changes on each diet differed significantly from each other: high-GI, -0·49 (SEM 0·29) KG; LOW-GI, -0·19 (sem 0·40) kg; MUFA +0·27 (sem 0·45) kg. Blood lipids did not change, but glycated haemoglobin increased significantly on MUFA, 0·02 (sem 0·11) %, relative to low-GI, -0·19 (sem 0·08) %, and high-GI, -0·13 (sem 0·14) %. Diastolic blood pressure fell by 8 mmHg on low-GI relative to MUFA (P=0·038). Although SI and AIRglu did not change significantly, DI, a measure of the ability of β-cells to overcome insulin resistance by increasing insulin secretion, increased on low-GI by >50 % (P=0·02). After adjusting for baseline values, the increase in DI on low-GI, 0·17 (sem 0·07), was significantly greater than those on MUFA, -0·09 (sem 0·08) and high-GI, -0·03 (sem 0·02) (P=0·019). Thus, the long-term effects of altering the source of dietary carbohydrate differ from those of altering the amount. High-carbohydrate–low-GI dietary advice improved β-cell function in subjects with IGT, and may, therefore, be useful in the management of IGT.

Elevated concentrations of fasting and non-fasting triacylglycerol-rich lipoproteins (TRL) as well as oxidative changes of lipoproteins may increase the risk of ischaemic heart disease. To compare the effects of different diets rich in unsaturated fatty acids on the concentrations and in vitro oxidation of fasting and postprandial lipoproteins eighteen males consumed diets enriched with rapeseed oil (RO), olive oil (OO), or sunflower-seed oil (SO) in randomised order for periods of 3 weeks followed by a RO test meal. In the postprandial state the concentrations of cholesterol and triacylglycerol (TAG) in TRL were higher after consumption of OO compared with RO and SO (P<0·04), possibly related to differences in the fasting state. The propagation rates for VLDL and LDL oxidation were higher in the postprandial compared with the fasting state irrespective of diet. In the fasting state, the propagation rates were highest after SO (P<0·001), and in the postprandial state, SO gave rise to a shorter VLDL lag time (P=0·03) and a higher propagation rate than OO consumption (P=0·04). Overall, the SO diet resulted in a higher postprandial propagation rate of LDL (P<0·001) compared with RO and OO, while there was no effect of diet on LDL oxidation lag time. Our results suggest that RO and SO diets lower the postprandial cholesterol and TAG concentrations compared with OO, while RO and OO diets result in similar and lower in vitro susceptibility to oxidation of lipoproteins than SO.

The effect of regular intake of low doses of an effervescent multivitamin preparation on the free-radical-producing activity of murine peritoneal macrophages under conditions resembling a possible infection was studied in vitro. Initially, several groups of mice were fed a basal diet and given, for 2 weeks, water without or with supplementation of either α -tocopherol, ascorbic acid, riboflavin or a multivitamin preparation. The supplementation period was followed by a 2-week wash-out time interval during which control and multivitamin groups received deionized water. Macrophage stimulation tests using chemiluminescent spectroscopy were performed at the end of the supplementation and wash-out periods to determine cell counts and their capacity to produce free radicals. Multivitamin supplementation increased the number, and the reactive oxygen species-producing activity, of macrophages. This effect persisted for 2 weeks after higher doses of supplementation were stopped. Multivitamin supplementation lowered the steady-state free radical concentrations of liver and spleen as measured by electron paramagnetic resonance spectroscopy. It also increased the antioxidant reactivity of the same organs, while there was no effect on the free radical concentration and antioxidant capacity of the kidney and brain. When taken regularly, low doses of multivitamin supplementation may have a beneficial effect on the defence mechanisms of the organism.

Pregnancy and diabetes lead to metabolic alterations in the energy balance that may not be completely independent. The objective of the present study was to look at the alterations induced by type 1 diabetes mellitus on the energy balance of pregnant rats and the offspring. Diabetes was induced by streptozotocin injection 15 d before the starting of pregnancy. The rats had their energy balance variables followed for 21 d. Protein, fat and energy content of dams was determined from samples of the carcasses. Pregnancy led to increased energy intake, energy gain and energy expenditure as well as higher gross food efficiency than non-pregnant counterparts. Diabetes increased metabolizable energy intake but not the energy gain of the animals: they had very high energy expenditure, so that diabetes blocked the improvement in gross food efficiency shown during pregnancy. Offspring from diabetic dams were born with lower body weight. Pregnant animals did not present the usual energy storage as seen by lower energy gain of diabetic dams as well as by the lower fat content in the carcasses of pregnant diabetic rats. It is concluded that diabetes impairs the energy variables usually enhanced by pregnancy alone.

The present study tested the hypothesis that habitual exercisers demonstrate an increased accuracy of regulation of food intake in compensation for previous dietary energy intake. Twenty-three lean healthy male subjects were divided into two groups on the basis of their habitual exercise levels: non-exercisers (no exercise sessions/week, n 9), and exercisers (>two exercise sessions of 40 min or more/week, n 14). The appetite response to covert liquid preloads of high (2513 kJ) energy (HE) and low (1008 kJ) energy (LE) was investigated Sixty minutes after the preload subjects were offered an ab libitum buffet-style meal and energy intake (EI) was calculated. Subjective hunger and satiety were assessed throughout using self-rated visual-analogue scales. Buffet EI in non-exercisers was not significantly different following the LE or HE preloads (mean compensation 7 %), but the exercise group significantly reduced their energy intake following the HE, compared with the LE, preload (mean compensation 90 %; P=0·0035). A broadly similar pattern of response was observed for both moderate (two to three sessions/week, n 7) and high exercisers (>four sessions/week, n 7). There were no significant differences between hunger or satiety ratings following HE or LE preloads for either group. However non-exercisers scored significantly higher on their self-ratings of hunger at the start of the study, before preload consumption, compared with the exercisers (P<0·01). These findings demonstrate that habitual exercisers have an increased accuracy of short-term regulation of food intake in compensation for preload manipulation, and provide additional support for advocating regular exercise in the prevention of overweight and obesity.